Cancer-associated fibroblasts (CAFs), a major component of tumor stroma, play an important role in tumor progression and metastasis via paracrine signaling, extracellular matrix (ECM) remodelling, and the secretion of growth factors, cytokines, and chemokines promoting tumor growth. In this study, we aim to develop a multitargeted nanoparticle formulation to simultaneously target CAFs and triple negative breast cancer (TNBC) cells, aiming to block their crosstalk, remodel the ECM, and inhibit epithelial-mesenchymal transition (EMT) to suppress tumor growth and metastatic processes. The potential of CAFs as therapeutic target for TNBC was evaluated by analyzing RNA-seq data from TNBC patients. A novel iRGD-functionalized polymer-lipid hybrid nanoparticle (iRGD-DOX-oHA-PLN) was specifically designed to target αvβ3 and αvβ5 receptors on both tumor cells and CAFs. A murine TNBC cell line and its orthotopic in vivo model were used for efficacy evaluation. Within this model, a murine fibroblast cell line was incorporated as a source of CAFs for in vitro investigation of the crosstalk between CAFs and tumor cells. In addition, the mechanism of iRGD-DOX-oHA-PLN nanoparticles in disrupting CAF-tumor cell interactions by blocking downstream features associated with metastases including paracrine communication, collagen organization and epithelial-mesenchymal transition (EMT) associated with metastasis were investigated both in vitro and in vivo. The iRGD-DOX-oHA-PLN demonstrates higher cellular uptake and cytotoxicity in CAFs overexpressing integrins αvβ3 and αvβ5 compared to normal fibroblasts. The results demonstrate that iRGD-DOX-oHA-PLN significantly restricts the interaction of CAFs and cancer cells, due to interruption of paracrine communication induced by pro-metastatic TGF-β and CXCL12 secretion together with a reduction of CXCR4 expression. Consistent with the in vitro results, in an orthotopic syngeneic TNBC murine model, iRGD-DOX-oHA-PLN effectively depletes CAFs, reducing tumor-associated collagen production, and altering tumor-associated collagen signatures (TACS). Thus, iRGD-DOX-oHA-PLN treatment impedes tumor invasion by reducing MMP-9 secretion and inhibiting EMT. The data reveals that simultaneously targeting CAFs and tumor cells, while blocking their crosstalk via application of a multitargeted nanomedicine, offers a compelling and effective strategy to inhibit metastasis of TNBC.

BACKGROUND:Up to date, Alzheimer's disease (AD) has very limited disease-modifying treatment. With respect to neuroprotection, brain-derived neurotrophic factor (BDNF) has been shown to promote the survival and synaptic plasticity of glutamatergic and GABAergic neurons in brain regions associated with cognitive and emotive decline relevant to AD. However, the poor blood-brain barrier (BBB) permeability and pharmacokinetic properties of BDNF limit its utilization as a neuroprotective treatment. Herein we aim to design a new BDNF nanocarrier system using a novel BBB-permeable terpolymer (BDNF-TPN) and investigate its neuroprotective effect in neurones and AD mice. METHOD:Bioactivity of the BDNF-TPN was first evaluated in vitro using the SH-SY5Y cell differentiation assay. The protective effects of BDNF-TPN on primary murine hippocampal neurons were assessed following exposure to Aβ. The BDNF delivery and expression in the brain following IV injection were examined by ELISA and confocal microscopy. The biodistribution and safety were evaluated via hematologic, clinical biochemical, immunotoxicity and histology tests in APP transgenic TgCRND8 AD mice and CD-1 mice. The effects of the treatment were evaluated in AD mice via immunohistochemistry, ELISA and behavioral test after 4-week IV treatment (weekly, 1 mg BDNF/kg b.w.). RESULT:The BDNF-TPN maintained BDNF bioactivity and rescued Aβ42 toxified primary neurons in vitro. Biomarker studies demonstrated BDNF-mediated neuroprotective signaling in transgenic mice following IV treatment using BDNF-TPN. Compared to free BDNF, BDNF-TPN significantly reduced reactive microglia and astrocytes and apoptosis of neurons. The pAKT level increased more than 2-fold in BDNF-TPN-treated group compared to vehicle and free BDNF treated groups. Synaptophysin, a marker for synaptic plasticity and integrity, was profoundly increased. The improved hippocampal-dependent contextual learning in the AD mice was observed. There was not detectable toxicity following the BDNF-TPN treatment. CONCLUSION:Our findings suggest BDNF-TPN is a promising treatment for reducing neuroinflammation, apoptosis and programmed cell death in AD mouse brains, while improving synaptic plasticity and cognitive function. Reference 1. Zhang W, et al. Sig Transduct Target Ther 2023, 8, 267. 2. He C, et al. Nano Today. 2020;35:100965. 3. Park E, et al. Advanced Science. 2023, 10(12):2207238. 4. Park E, et al. Biomaterials. 2025 Jan 24:123142.

Alzheimer's disease (AD) affects over 50 million people worldwide and is the sixth leading cause of death in North America with no cure and limited disease-modifying therapies. Brain-derived neurotrophic factor (BDNF) is a potential therapeutic agent, as it promotes the survival and synaptic plasticity of glutamatergic and GABAergic neurons in brain regions associated with cognitive and emotive decline in AD, and mice lacking BDNF exhibits hippocampal long-term potentiation (LTP) impairment, reversible with recombinant BDNF. However, the poor blood-brain barrier (BBB) permeability and pharmacokinetic properties of BDNF limit its clinical use. To address these limitations, a BDNF nanocarrier system is designed using a novel BBB-permeable terpolymer (TP). This terpolymer-based nanoparticle system (BDNF-TPN) with optimized physiochemical properties protects BDNF in circulation and enables sufficient neuronal delivery of BDNF and neuron survival following Aβ exposure in vitro. The biodistribution and safety of BDNF-TPN are evaluated via imaging, enzyme-linked immunosorbent assay, hematologic, clinical biochemical, immunotoxicity, and histology tests. Biomarker studies demonstrate BDNF-mediated neuroprotective signaling in an APP transgenic mouse model following intravenous injection of BDNF-TPN with reduced apoptosis and neuron inflammation and increased neuronal survival and synaptic plasticity. Improved hippocampal-dependent contextual learning in APP transgenic Alzheimer's mice is observed as determined by fear-conditioning assay following two months of weekly treatments.

Triple negative breast cancer (TNBC) is an aggressive malignancy characterized by early recurrence, high metastatic burden, and resistance to conventional therapies, largely due to the absence of targetable receptors and an immunosuppressive tumor microenvironment. To address these limitations, our lab has engineered a multifunctional nanotherapeutic system iRGD-DOX-oHA-PLN comprising polymer-lipid nanoparticles co-loaded with doxorubicin (DOX) and oligomeric hyaluronic acid (oHA) and functionalized with the tumor-penetrating integrin-targeting iRGD peptide. This rationally designed platform capitalizes on the sequential targeting mechanism: integrin-mediated tumor penetration and endocytosis via iRGD, followed by CD44 engagement through oHA to enhance intracellular drug delivery and suppresses cell motility. iRGD-DOX-oHA-PLN significantly improved cellular uptake, intratumoral accumulation, and cytotoxic efficacy in TNBC cells and tumors. Notably, it enhanced immunogenic cell death, characterized by increased calreticulin exposure, ATP and HMGB1 levels, triggering potent anti-tumor immune responses. Intravenous treatment led to elevated CD8+ T-cell infiltration, granzyme B expression, and secretion of pro-inflammatory cytokines (TNF-α, IFN-γ), while concurrently suppressing immunosuppressive mediators including IL-6, regulatory T cells, and tumor-associated macrophages. Over a four-week treatment period, iRGD-DOX-oHA-PLN effectively inhibited primary tumor growth and systemic pulmonary metastases in a syngeneic orthotopic TNBC mouse model. These findings demonstrate the therapeutic potential of simultaneously targeting integrin-CD44 signaling and the immunosuppressive niche using a dual-functional nanomedicine to overcome drug resistance and immune evasion in TNBC, offering a promising strategy for metastatic cancer intervention.

The development of disease-modifying therapeutics for Alzheimer's disease remains challenging due to the complex pathology and the presence of the blood-brain barrier. Previously we have described the investigation of a brain-penetrating multifunctional bioreactive nanoparticle system capable of remodeling the hypoxic and inflammatory brain microenvironment and reducing beta-amyloid plaques improving cognitive function in a mouse model of Alzheimer's disease. Despite the linkage of hypoxia and inflammation to metabolic alteration, the effects of this system on modulating cerebral glucose metabolism, mitochondrial activity and synaptic function remained to be elucidated. To examine this, a transgenic mouse model of Alzheimer's disease (TgCRND8) in vivo were treated intravenously with beta-amyloid antibody-conjugated (Ab), blood-brain barrier-crossing terpolymer (TP) containing polymer-lipid based manganese dioxide nanoparticles (Ab-TP-MDNPs). Alterations in cerebral glucose utilization were determined by [1⁸F]FDG-PET imaging in vivo, with glucose metabolism and mitochondrial activity analyzed by biomarkers and studies with primary neurons in vitro. Synaptic function was evaluated by both biomarkers and electrophysiologic analysis. Current study shows that intravenously administered Ab-TP-MDNPs enhanced cerebral glucose utilization, improved glucose metabolism, mitochondrial activity, and increased the levels of neprilysin, O-glycosylation. The consequence of this was enhanced glucose and ATP availability, resulting in improved long-term potentiation for promoting neuronal synaptic function. This study highlights the importance of targeting the metabolism of complex disease pathologies in addressing disease-modifying therapeutics for neurodegenerative disorders such as Alzheimer's disease.

Alzheimer's disease (AD), a progressive neurodegenerative disorder, is closely associated with the aggregation of amyloid-beta (Aβ) peptides, particularly Aβ42. Toxic oligomeric forms of Aβ42 are implicated in synaptic dysfunction and cognitive decline, making them a key target for therapeutic interventions 1 . This study explores the interactions between a novel biodegradable terpolymer we developed for delivering imaging and therapeutic agents to the AD brain 2,3 and Aβ42 to evaluate its therapeutic potential. The biding affinity between Aβ42 and the terpolymer was evaluated using biolayer interferometry (BLI). Replica exchange molecular dynamics (REMD) 4 simulation was performed to analyze conformational changes in Aβ42 upon binding to the terpolymer. Circular dichroism (CD) spectroscopy, transmission electron microscopy (TEM), and confocal microscopy were employed to monitor time course of structural transitions, aggregation patterns, and cellular interactions. SH-SY5Y cells were used to assess the effect of the terpolymer on attenuating Aβ42-induced cytotoxicity. REMD simulations revealed rapid conformational transitions in Aβ42 upon terpolymer binding, shifting from random coil and α-helical structures to stabilized β-sheet-rich states. CD spectroscopy demonstrated accelerated secondary structure transitions, corroborating the simulation findings. TEM showed that the terpolymer disrupted conventional Aβ42 aggregation patterns, resulting in smaller, irregular aggregates rather than typical long fibrils. Cellular assays indicated that the terpolymer significantly reduced Aβ42-induced cytotoxicity, improving cell viability in the terpolymer treated groups. Confocal microscopy revealed cellular uptake of Aβ42-terpolymer complexes, reducing extracellular toxicity and harmful interactions with cellular membranes. This study highlights the capability of the terpolymer in modulating Aβ42 aggregation dynamics, mitigating toxic oligomer formation and alleviating its neurotoxic effects. These findings present a promising approach for developing polymer-based therapies targeting Alzheimer's disease, offering a novel pathway to combat neurodegeneration. References Okumura, H., J Phys Chem B 2023, 127 (51), 10931-10940. Park E, et al. Advanced Science. 2023, 10(12):2207238. Li, L.Y.; Park, E. et al. Nanotoxicology 2024 , 1-20. Li, L.Y.; Park, E. et al. Nanotoxicology 2024, 1-20.

Up to date, Alzheimer's disease (AD) has very limited disease-modifying treatment. With respect to neuroprotection, brain-derived neurotrophic factor (BDNF) has been shown to promote the survival and synaptic plasticity of glutamatergic and GABAergic neurons in brain regions associated with cognitive and emotive decline relevant to AD. However, the poor blood-brain barrier (BBB) permeability and pharmacokinetic properties of BDNF limit its utilization as a neuroprotective treatment. Herein we aim to design a new BDNF nanocarrier system using a novel BBB-permeable terpolymer (BDNF-TPN) and investigate its neuroprotective effect in neurones and AD mice. Bioactivity of the BDNF-TPN was first evaluated in vitro using the SH-SY5Y cell differentiation assay. The protective effects of BDNF-TPN on primary murine hippocampal neurons were assessed following exposure to Aβ. The BDNF delivery and expression in the brain following IV injection were examined by ELISA and confocal microscopy. The biodistribution and safety were evaluated via hematologic, clinical biochemical, immunotoxicity and histology tests in APP transgenic TgCRND8 AD mice and CD-1 mice. The effects of the treatment were evaluated in AD mice via immunohistochemistry, ELISA and behavioral test after 4-week IV treatment (weekly, 1 mg BDNF/kg b.w.). The BDNF-TPN maintained BDNF bioactivity and rescued Aβ42 toxified primary neurons in vitro. Biomarker studies demonstrated BDNF-mediated neuroprotective signaling in transgenic mice following IV treatment using BDNF-TPN. Compared to free BDNF, BDNF-TPN significantly reduced reactive microglia and astrocytes and apoptosis of neurons. The pAKT level increased more than 2-fold in BDNF-TPN-treated group compared to vehicle and free BDNF treated groups. Synaptophysin, a marker for synaptic plasticity and integrity, was profoundly increased. The improved hippocampal-dependent contextual learning in the AD mice was observed. There was not detectable toxicity following the BDNF-TPN treatment. Our findings suggest BDNF-TPN is a promising treatment for reducing neuroinflammation, apoptosis and programmed cell death in AD mouse brains, while improving synaptic plasticity and cognitive function. Reference 1. Zhang W, et al. Sig Transduct Target Ther 2023, 8, 267. 2. He C, et al. Nano Today . 2020;35:100965. 3. Park E, et al. Advanced Science . 2023, 10(12):2207238. 4. Park E, et al. Biomaterials . 2025 Jan 24:123142.

Neuroinflammation plays a causal role in neurodegenerative Alzheimer’s disease (AD); it occurs long before clinical onset of AD. 1 Therefore, early detection of neuroinflammation is critical for early intervention before the irreversible neurodegeneration happens. To address this pressing need, our group has developed multifunctional bioreactive nanoparticles, consisting of blood-brain barrier-penetrating terpolymer and MnO 2 nanoparticles and conjugated anti-Ab antibody (Ab-TP-MDNP). The system reduced oxidative stress and produced oxygen and paramagnetic Mn 2+ ions, thereby remodeling the brain microenvironment and enabling sensitive detection of early neuroinflammation prior to Ab plague formation in an APP transgenic TgCRND8+ AD mouse model. 2 We also demonstrated its effects on improving vascular functions, Ab elimination, energy metabolism, neuronal activity and cognitive function. 3-4 Built on the foundation of previous findings, we investigate whether the TP-MDNP is able enhance early detection of neuroinflammation regardless of Aß or tau expression and evaluate its therapeutic effect in AD mouse model of tauopathy. Three types of transgenic mouse model of AD, TgCRND8+, PS19 with tauopathy, and APP/PS1 were used in the MRI study. The diagnostic performance of Ab-TP-MDNP in MRI was also compared with PET imaging using F18-florobetaben in TgCRND8+ mice. PS19 mice were treated with IV injection of TP-MDNP for two weeks (2/week, 100 μmol Mn/kg b.w.) and the biomarkers for ROS, neuroinflammation, and p-Tau expression were examined using immunohistochemistry and ELISA. Ab-TP-MDNP enhanced MRI signal significantly outperformed PET imaging by Ab-targeted F18-florobetaben in TgCRND8+ mice of 3 months and 6 months of age. Similar MRI imaging sensitivity was observed in PS19 and APP/PS1 mice with or without conjugated antibody against Ab or tau protein, suggesting the neuroinflammation activated MRI contrast enhancement as a common mechanism. In PS19, TP-MDNP treatment significantly reduced total ROS, CA9 (a hypoxia marker) and p-tau levels. The results suggest that TP-MDNP can enable MRI detection of neuroinflammation and reduce neurodegeneration pathogenetic factors such as ROS, hypoxia, and p-tau in AD mouse brains. References 1. Zhang W, et al. Sig Transduct Target Ther 2023;8, 267. 2. He C, et al. Nano Today . 2020;35:100965. 3. Park E,. et al. Advanced Science . 2023 Apr;10(12):2207238. 4. Park E, et al. Biomaterials . 2025 Jan 24:123142.

The high mortality rate associated with metastatic breast cancer presents a significant global challenge. Inherent and chemotherapy-induced DNA damage repair, alongside immunosuppression, drastically contribute to triple-negative breast cancer (TNBC) relapse and metastasis. While poly (ADP-ribose) polymerase (PARP) inhibitors such as olaparib show effectiveness against BRCA1-mutant TNBC, they may lead to drug resistance and reduced efficacy due to increased programmed death-ligand 1 (PD-L1) expression. Our study explored the use of polymer-lipid nanoparticles (PLN) loaded with doxorubicin (DOX) and oligomeric hyaluronic acid (oHA), functionalized iRGD-peptide for integrins targeting (iRGD-DOX-oHA-PLN), to prevent TNBC immunosuppression, DNA repair, and metastasis. The results demonstrate that the iRGD-DOX-oHA-PLNs efficiently downregulated single and double-strand DNA repair proteins and enhanced DNA damage while decreasing PD-L1 expression compared to olaparib. Accordingly, iRGD-DOX-oHA-PLN treatment showed significantly higher efficiency in reducing levels of primary tumor growth and numbers of metastases to the lung and liver compared to olaparib in vitro and in vivo in both BRCA1-mutant and wild type TNBC orthotopic xenograft models.

A novel brain-targeted and reactive oxygen species-activatable manganese dioxide containing nanoparticle system functionalized with anti-amyloid-β antibody (named aAβ-BTRA-NC) developed by our group has shown great promise as a highly selective magnetic resonance imaging (MRI) contrast agent for early detection and multitargeted disease-modifying treatment of Alzheimer's disease (AD). To further evaluate the suitability of the formulation for future clinical application, we investigated the safety, biodistribution, and pharmacokinetic profile of aAβ-BTRA-NC in a transgenic TgCRND8 mouse AD model, wild type (WT) littermate, and CD-1 mice. Dose-ascending studies demonstrated that aAβ-BTRA-NC was well-tolerated by the animals up to 300 μmol Mn/kg body weight [b.w.], 3 times the efficacious dose for early AD detection without apparent adverse effects; Histopathological, hematological, and biochemical analyses indicated that a single dose of aAβ-BTRA-NC did not cause any toxicity in major organs. Immunotoxicity data showed that aAβ-BTRA-NC was safer than commercially available gadolinium-based MRI contrast agents at an equivalent dose of 100 μmol/kg b.w. of metal ions. Intravenously administered aAβ-BTRA-NC was taken up by main organs with the order of liver, kidneys, intestines, spleen, followed by other organs, and cleared after one day to one week post injection. Pharmacokinetic analysis indicated that the plasma concentration profile of aAβ-BTRA-NC followed a 2-compartmental model with faster clearance in the AD mice than in the WT mice. The results suggest that aAβ-BTRA-NC exhibits a strong safety profile as a nanotheranostic agent which warrants more robust preclinical development for future clinical applications.

Radiotherapy (RT) is one of major therapeutic modalities in combating breast cancer. In RT, ionizing radiation is employed to induce DNA double-strand breaks (DSBs) as a primary mechanism that causes cancer cell death. However, the induced DNA damage can also trigger the activation of DNA repair mechanisms, reducing the efficacy of RT treatment. Given the pivotal role of RAD50 protein in the radiation-responsive DNA repair pathways involving DSBs, we developed a novel polymer-lipid based nanoparticle formulation containing RAD50-silencing RNA (RAD50-siRNA-NPs) and evaluated its effect on the RAD50 downregulation as well as cellular and tumoral responses to ionizing radiation using human triple-negative breast cancer as a model. The RAD50-siRNA-NPs successfully preserved the activity of the siRNA, facilitated its internalization by cancer cells via endocytosis, and enabled its lysosomal escape. The nanoparticles significantly reduced RAD50 expression, whereas RT alone strongly increased RAD50 levels at 24 h. Pretreatment with RAD50-siRNA-NPs sensitized the cancer cells to RT with ∼2-fold higher level of initial DNA DSBs as determined by a γH2AX biomarker and a 2.5-fold lower radiation dose to achieve 50% colony reduction. Intratumoral administration of RAD50-siRNA-NPs led to a remarkable 53% knockdown in RAD50. The pretreatment with RAD50-siRNA-NPs followed by RT resulted in approximately a 2-fold increase in DNA DSBs, a 4.5-fold increase in cancer cell apoptosis, and 2.5-fold increase in tumor growth inhibition compared to RT alone. The results of this work demonstrate that RAD50 silencing by RAD50-siRNA-NPs can disrupt RT-induced DNA damage repair mechanisms, thereby significantly enhancing the radiation sensitivity of TNBC MDA-MB-231 cells in vitro and in orthotopic tumors as measured by colony forming and tumor regrowth assays, respectively.

Finding effective disease-modifying treatment for Alzheimer's disease remains challenging due to an array of factors contributing to the loss of neural function. The current study demonstrates a new strategy, using multitargeted bioactive nanoparticles to modify the brain microenvironment to achieve therapeutic benefits in a well-characterized mouse model of Alzheimer's disease. The application of brain-penetrating manganese dioxide nanoparticles significantly reduces hypoxia, neuroinflammation, and oxidative stress; ultimately reducing levels of amyloid β plaques within the neocortex. Analyses of molecular biomarkers and magnetic resonance imaging-based functional studies indicate that these effects improve microvessel integrity, cerebral blood flow, and cerebral lymphatic clearance of amyloid β. These changes collectively shift the brain microenvironment toward conditions more favorable to continued neural function as demonstrated by improved cognitive function following treatment. Such multimodal disease-modifying treatment may bridge critical gaps in the therapeutic treatment of neurodegenerative disease.

Despite substantial progress in the treatment of castration-resistant prostate cancer (CRPC), including radiation therapy and immunotherapy alone or in combination, the response to treatment remains poor due to the hypoxic and immunosuppressive nature of the tumor microenvironment. Herein, we exploited the bioreactivity of novel polymer–lipid manganese dioxide nanoparticles (PLMDs) to remodel the tumor immune microenvironment (TIME) by increasing the local oxygen levels and extracellular pH and enhancing radiation-induced immunogenic cell death. This study demonstrated that PLMD treatment sensitized hypoxic human and murine CRPC cells to radiation, significantly increasing radiation-induced DNA double-strand breaks and ultimately cell death, which enhanced the secretion of damage-associated molecular patterns, attributable to the induction of autophagy and endoplasmic reticulum stress. Reoxygenation via PLMDs also polarized hypoxic murine RAW264.7 macrophages toward the M1 phenotype, enhancing tumor necrosis factor alpha release, and thus reducing the viability of murine CRPC TRAMP-C2 cells. In a syngeneic TRAMP-C2 tumor model, intravenous injection of PLMDs suppressed, while radiation alone enhanced recruitment of regulatory T cells and myeloid-derived suppressor cells. Pretreatment with PLMDs followed by radiation down-regulated programmed death-ligand 1 and promoted the infiltration of antitumor CD8 + T cells and M1 macrophages to tumor sites. Taken together, TIME modulation by PLMDs plus radiation profoundly delayed tumor growth and prolonged median survival compared with radiation alone. These results suggest that PLMDs plus radiation is a promising treatment modality for improving therapeutic efficacy in radioresistant and immunosuppressive solid tumors.

Background: Metastatic triple-negative breast cancer (TNBC), especially with BRCA1/2 mutations, is a deadly subtype of breast cancer with ~12% of 5-year survival. Despite their FDA approval, the therapeutic benefits of poly(adenosine diphosphate-ribose) polymerase inhibitors (PARPi) are limited to BRCA1/2 mutated malignancies. Moreover, the PARPi olaparib has been shown to upregulate the expression of programmed death-ligand 1 (PD-L1) leading to immune suppression. The purpose of this study is to investigate if a novel nanoparticle formulation of oligomer hyaluronic acid (oHA) and doxorubicin (DOX) co-loaded in an iRGD-conjugated polymer-lipid nanocarrier (iRGD-DOX-oHA-PLN) can inhibit DNA damage repair and suppress PD-L1 in both mutant and non-mutant TNBC cells and reduce tumor progression and lung metastasis compared to the PARPi olaparib. We postulate that the co-administered oHA will block the signaling pathways of native hyaluronic acid (HA) receptors, the cluster of differentiation 44 (CD 44) and receptor for HA mediated motility (RHAMM), that regulate DNA damage repair and immunosuppression and enhance DOX efficacy. Methods: In vitro cellular uptake of various DOX formulations by TNBC MDA-MB-231-luc-D3H2LN cells and MDA-MB-436 (BRCA1 mutant) cells was examined using confocal laser scanning microscopy or spectrophotometer. The expression level of RHAMM, PD-L1, and PARP1 parylation (PAR) was evaluated after treatment with iRGD-DOX-oHA-PLN or olaparib in both cell lines using western blot or confocal laser scanning microscopy. The therapeutic efficacy of iRGD-DOX-oHA-PLN compared to olaparib was determined by monitoring the tumor progression and lung metastasis development in orthotopic breast tumor models of MDA-MB-231-luc-D3H2LN or MDA-MB-436 cell line. Results: The expression of native HA receptor RHAMM was profoundly reduced by ~5 folds in vivo and the DNA DSB was significantly increased by the iRGD-DOX-oHA-PLN treatment with 40% of γH2AX positive cells in tumor tissue sections. The iRGD-functionalized PLN enhanced DOX cellular uptake compared to DOX free drug or non-targeted NPs by ~3- and ~1.5-fold, in MDA-MB-231 and MDA-MB-436 cells, respectively. While olaparib upregulated PD-L1 and free DOX increased PAR level, the iRGD-DOX-oHA-PLN reduced the parylation and PD-L1 expression in both TNBC cell lines. The iRGD-DOX-oHA-PLN treatment (two dose of 10 mg/kg i.v. biweekly) outperformed olaparib (twenty doses of 50 mg/kg i.p 5x/week) in preventing tumor progression and lung metastasis in vivo over a 4-week period. Conclusion: The results suggest that the iRGD-DOX-oHA-PLN can effectively inhibit DNA damage repair and immunosuppression of cancer cells and could be a promising multitargeted nanomedicine for the treatment of both BRCA1-mutant and non-mutant metastatic TNBC. Citation Format: Ibrahim Alradwan, Pei Zhi, Tian Zhang, HoYin Lip, Abdulmutalib Zetrini, Chunsheng He, Jeffery Henderson, Andrew Rauth, Xiao Yu Wu. Targeted nanoparticles reduce tumor progression and lung metastasis by limiting DNA damage repair and immune suppression in metastatic triple-negative breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5990.

Globally, a rising burden of complex diseases takesa heavy toll on human lives and poses substantial clinical andeconomic challenges. This review covers nanomedicine andnanotechnology-enabled advanced drug delivery systems (DDS)designed to address various unmet medical needs. Key nano-medicine and DDSs, currently employed in the clinic to tacklesome of these diseases, are discussed focusing on their versatility indiagnostics, anticancer therapy, and diabetes management. First-hand experiences from our own laboratory and the work of othersare presented to provide insights into strategies to design andoptimize nanomedicine- and nanotechnology-enabled DDS forenhancing therapeutic outcomes. Computational analysis is alsobriefly reviewed as a technology for rational design of controlledrelease DDS. Further explorations of DDS have illuminated the interplay of physiological barriers and their impact on DDS. It isdemonstrated how such delivery systems can overcome these barriers for enhanced therapeutic efficacy and how new perspectives ofnext-generation DDS can be applied clinically.

Background: Triple-negative breast cancer (TNBC) is a highly aggressive breast cancer subtype. Despite the early response to chemotherapy, high incidence of recurrence leads to short overall survival and poor prognosis. Native hyaluronic acid (HA) interacts with CD44 and receptors for hyaluronan mediated motility (RHAMM) overexpressed in TNBC mediating chemo-resistance and metastasis. In contrast, oligomeric HA (oHA) can disrupt HA-CD44/RHAMM interactions and attenuate oncogenic and pro-metastatic pathways, such as the mitogen-activated protein kinase/extracellular-signal-regulated kinase (MAPK/ERK) pathway, reducing downstream DNA repair and drug resistance markers. Our previous work demonstrated that oHA can exert synergistic anti-tumor and anti-metastatic effects when combined with doxorubicin (DOX) and co-loaded in an integrin-targeted iRGD-modified nanoparticle system (iRGD-DOX-oHA-PLNs). This study aims to investigate the inhibitory effect of iRGD-DOX-oHA-PLNs on both DNA single-strand break (SSB) and double-strand break (DSB) repair proteins and drug efflux pumps responsible for multidrug resistance to enhance DOX efficacy in breast cancer gene 1 (BRCA1) mutant and non-mutant TNBC. Methods: The cytotoxicity of DOX, oHA and their combinations in free solution or in nanoparticles was evaluated by clonogenic assay in human MDA-MB-231-luc-D3H2LN and MDA-MB-436 (BRCA1 mutant) TNBC cells. The in vitro expression of a DNA DSB marker, DNA repair markers, and drug efflux pump P-glycoprotein (P-gp) were measured by Western blot. The in vivo expression level of BRCA1 and Rad51 in an orthotopic TNBC mouse model was determined by immunohistochemical staining. Results: The combination of DOX-oHA showed synergism against both BRCA1 mutant and non-mutant TNBC cells. The iRGD-DOX-oHA-PLNs induced great increases in DNA DSBs demonstrated by the highest γH2AX level compared to other treatment groups. These nanoparticles also showed inhibitory effects on the expression of both DNA SSB repair protein (poly (ADP-ribose) polymerase) and DNA DSB repair proteins (Rad50 and Rad51), contributing to the enhanced efficacy of chemotherapy. The immunohistochemical staining of tumor tissues indicated lower levels of BRCA1 and Rad51 after iRGD-DOX-oHA-PLN treatment than the formulation without oHA, attributable to the effect of intracellularly delivered oHA on limiting the MAPK signaling. Additionally, iRGD-DOX-oHA-PLNs reduced the expression of the drug efflux pump P-gp as compared to DOX treatment groups without oHA. Conclusion: The co-delivery of oHA and DOX in the iRGD-DOX-oHA-PLNs efficiently blocked DNA damage repair and down-regulated the drug efflux pump P-gp, thus improving the efficacy of DOX. Collectively, this nanoparticle system could be a promising option for metastatic TNBC treatment. Citation Format: Pei Zhi, Ibrahim Alradwan, Tian Zhang, HoYin Lip, Abdulmutalib Zetrini, Chunsheng He, Jeffery Henderson, Andrew Michael Rauth, Xiao Yu Wu. Synergistic combination nanomedicine of doxorubicin and oligo hyaluronic acid inhibits DNA damage repair and overcomes drug resistance in metastatic triple-negative breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5399.

Developing effective disease-modifying treatment for Alzheimer’s disease (AD) remains a tremendous challenge due to its multifactorial nature involving multiple pathologic signaling pathways in addition to ineffective drug delivery through the blood-brain barrier (BBB). 1 With this in mind our group has developed multifunctional bioreactive nanoparticles (Ab-TP-MDNPs), consisting of anti-amyloid β antibody (Ab) linked to brain-penetrating terpolymer (TP) and manganese dioxide (MnO 2 ) nanoparticles (MDNPs), that are shown to reduce oxidative stress in AD brains. 2 Given the early occurrence of oxidative stress, hypoxia, and vascular dysfunction in AD brains, 3,4 we investigated the therapeutic effects of Ab-TP-MDNPs on reducing neuroinflammation and vascular dysfunction in an AD mouse model. A transgenic mouse model of AD (TgCRND8 species) and wildtype littermates (WT) were treated with intravenous (i.v.) injection of Ab-TP-MDNPs (twice/week, 100 µmol Mn/kg b.w.) or vehicle for 2-weeks. Oxidative and inflammatory biomarkers were examined using immunohistochemistry and enzyme-linked immunosorbent assay (ELISA). Vascular function before and after the treatment was studied via high resolution magnetic resonance imaging (MRI). Cerebral blood flow (CBF) was assessed using FAIR (flow-sensitive alternating inversion recovery) technique. BBB permeability was measured via T1 mapping re-acquisition prior to and following i.v. injection of gadolinium-diethylenetriamine penta-acetate (Gd-DTPA) at 1.2 mmol/kg Ab-TP-MDNPs treatment significantly decreased inflammatory cytokines and activation of microglia and astrocytes markers (reactive microglia: hippocampus by 69% and cortex by 59%, reactive astrocytes: hippocampus by 32% and cortex by 33%). In addition, Ab-TP-MDNPs treatment improved CBF (cortex by 19% and subcortex by 35%) and vessel leakage by 29% in the cortex of AD mouse brains. Ab-TP-MDNPs treatment reduced neuroinflammation and vascular dysfunction in an AD mouse model. These findings suggest a new multimodal strategy for AD treatment and encourage further development of such approach for complex neurologic diseases. Reference: 1.Panza F, Lozupone M, Logroscino G, Imbimbo BP. Nature Reviews Neurology . 2019;15(2):73-88. 2.He C, Ahmed T, Abbasi AZ, et al. Nano Today . 2020;35:100965. 3.Sweeney MD, Montagne A, Sagare AP, et al. Alzheimer’s & Dementia . 2019;15(1):158-167. 4.Nortley R, Korte N, Izquierdo P, et al. Science . 2019:eaav9518.

Abstract Background: Magnetic resonance image-guided radiation therapy (MRgRT) is a new-generation approach to improving treatment outcomes by allowing real-time precise tumor delineation that guides the radiation. However, the existing gadolinium-based contrast agents (GBCAs) are insufficient for MRgRT due to their severe side effects and rapid clearance from the tumor that requires multiple injections to maintain the signal. In addition, tumor hypoxia induced radiation resistance, which attenuates the RT efficacy. To date, there is no FDA-approved contrast agent with high safety and efficacy profile exhibiting both MR signal enhancement and RT sensitization capabilities. Based on our previous findings that manganese-dioxide nanoparticles (MDNP) can convert tumoral ROS (H2O2) into O2 to reduce hypoxia and sensitize RT, we propose the design of a novel tumor-targeting manganese-dioxide nanoparticle (MDNP) with dual MRI and RT enhancement functionalities. In this work, we evaluate the safety profile, biodistribution, in vivo clearance, MR contrast enhancement, and radiation sensitization effect of the MDNPs. Methods: Terpolymer-based MDNP (T-MDNP) were prepared by loading MnO2 precursor in a polymer-lipid matrix. Physicochemical properties and stability of NPs were determined by TEM, DLS, and zeta potential measurements. In vitro cellular uptake by cancer cells was measured via confocal microscopy. The biocompatibility and safety of the NPs was examined in vitro and in vivo. Tissue distribution, clearance, and tumor retention of T-MDNPs was evaluated using MRI and ICP. The tumoral MR signal enhancement by T-MDNP as a function of time was measured and compared with Gadavist™ (gadobutrol) in murine tumor models with human MDA-MB-231 breast or prostate PC3 xenografts. The ability of NPs to modulate tumor microenvironment (TME) and enhance RT efficacy was also evaluated. Results: The prepared T-MDNPs were 120 nm in size and exhibited excellent storage stability at room temperature and 4°C. The NPs showed negligible hemolysis effect suggesting their suitability for intravenous (IV) injection. As a result of in situ generation of Mn+2 ions via the reaction with tumoral ROS, a single dose of 70 μmole Mn/kg of body weight of T-MDNP increased tumor T1 signal by up to 1.4-fold and maintained it up to 4 hrs, while Gadavist™ was eliminated in about 30 min. The NPs were completely cleared from major organs in 72 hrs, as determined by MRI and ICP. Combination of NP treatments with RT resulted in significantly increased median survival time by 2-fold in human PC3 prostate model and 5-fold in human breast tumor model. Conclusions: Our results have demonstrated that the T-MDNPs are safe and effective as a dual MR contrast agent and radiation sensitizer. This system is promising for precise tumor margin delineation, maintaining MR signal in tumor for a duration needed for MRgRT, and improving RT efficacy. These properties make it an excellent candidate and the first-in-class entity that can be utilized with MRgRT. Citation Format: Charles Yen, Azhar Z. Abbasi, Chunsheng He, Mohammad Ali Amini, Hoyin Lip, Michael Rauth, Xiao Yu Wu. Theragnostic tumor-targeted manganese dioxide-loaded polymer-lipid nanoparticles for magnetic resonance image-guided radiation therapy [abstract]. In: Proceedings of the AACR Virtual Special Conference on Radiation Science and Medicine; 2021 Mar 2-3. Philadelphia (PA): AACR; Clin Cancer Res 2021;27(8_Suppl):Abstract nr PO-100.